Comparison of sperm preparation methods: effect on chromatin and morphology recovery rates and their consequences on the clinical outcome after in vitro fertilization embryo transfer

Abstract

The aim of this study was to evaluate the ef®cacy of swim-up, PureSperm Ò gradient centrifugation and glass±wool ®ltration methods for semen preparation and to assess the possible enhancement of the quality of the subpopulation of spermatozoa in terms of sperm concentration, morphology and chromatin condensation. Moreover, to determine the effect of this semen processing technique on the clinical outcome after in vitro fertilization embryo transfer (IVF-ET).A total of 180 semen samples of patients' husbands who were undergoing IVF therapy were prepared by swim-up (G1, n 60), PureSperm Ò gradient centrifugation (G2, n 60) or glass±wool (G3, n 60) methods. Chromatin condensation was assessed by Chromomycin (CMA3), whereas sperm morphology was evaluated according to strict criteria. In all three semen processing methods, the percentage of chromatin condensed and morphologically normal spermatozoa was higher after semen processing in comparison with native semen samples. The proportion of normal chromatin condensed spermatozoa prepared in glass±wool ®ltration was signi®cantly higher than that in swim-up (G.I, p 0.02) or PureSperm Ò (G.II, p 0.001). In addition semen processing with PureSperm Ò yields signi®cantly a higher percentage of morphologically normal spermatozoa than swim-up (p < 0.001) or glass±wool method (p < 0.002). However, the fertilization, implantation and pregnancy rates, in turn were similar in all semen preparation methods.In conclusion, PureSperm Ò gradient centrifugation yields a higher percentage of morphologically normal spermatozoa than shown in traditional swim-up or glass±wool ®ltration. However, the percentage of chromatin condensed spermatozoa was signi®-cantly higher after semen processing via glass±wool in comparison with the other two methods.Nevertheless, there were no signi®cant difference in the fertilization, implantation and pregnancy rates of sperm prepared by means of swim-up, PureSperm Ò or glass±wool ®ltration. Therefore, glass±wool ®ltration should be recommended as the ®rst choice for semen preparation for Intracytoplasmic sperm injection (ICSI) technique as the natural selection is bypassed. Whereas, swim-up and PureSperm should be used for semen processing in IVF programme.

Cryopreservation and Sperm DNA Integrity

“…There are several reports confirming the results from this study on the superior performances of the swim-up procedure to improve the sperm nuclear integrity and the overall semen quality of the fresh unselected semen, basing the analysis of defective sperm either by using the SCSA (Golan et al 1997;Spano`et al 1999;Zini et al 2000) or other techniques (Molina et al 1995;Younglai et al 2001;Lachaud et al 2004). However, other studies reported different findings with other separation techniques being more efficient (Hammadeh et al 2001a;Sakkas et al 2000). Noteworthy, among the WHO parameter, motility exhibits the strongest correlation with the SCSA DFI (Giwercman et al 2003): this can explained on the basis that sperm acquire motility and in parallel complete the nuclear packaging during the epididymal transit.…”

“…Freezing of sperm before initiation of treatment gives patients a sort of fertility insurance that may allow them to father their own children through the use of IVF or intracytoplasmic sperm injection (ICSI). Despite many refinements in cryopreservation and separation methodology, the salvage of post-thaw sperm remains poor even though it seems not to hamper fertilization or pregnancy rate in ART (Hammadeh et al 2001a;Kuczynski et al 2001). The most commonly reported detrimental effect of cryopreservation on human sperm is a marked reduction in motility (Kramer et al 1993), but also DNA integrity has recently started to be extensively checked.…”

Sperm DNA damage measured by the alkaline Comet assay as an independent predictor of male infertility and in vitro fertilization success

“…A reduction in DNA fragmentation was also seen in all samples after DGC. This was probably due to the removal of senescent and defective sperm (24)(25)(26)(27)(28)(29)(30)(31)(32)(33)(34)(35)(36). However, it is in conflict with some reports (37)(38)(39)(40).…”

The ability of sperm selection techniques to remove single- or double-strand DNA damage

“…[18][19][20][21] In our study, sperm selected after DGC and SUP exhibited sperm subpopulations from which highly motile spermatozoa were collected from the overlying buffer. Similar results have been previously reported by Ricci et al 22 and Boomsma et al…”